Voltage-gated sodium channels are critical determinants of neuronal excitability. Experimental and clinical evidence clearly demonstrates that sodium channels and changes in the properties of these channels can play crucial roles in chronic pain and other neurological disorders. However, at least six different sodium channel isoforms are expressed in peripheral sensory neurons, and investigating the role of specific voltage- gated sodium channel isoforms in normal and abnormal electrical activity of sensory neurons has been hindered by the lack of isoform specific neuronal sodium channel blockers. The goal of this R21 grant proposal is to develop selective sodium channel blockers based on biological peptide toxins that can be used to investigate the role(s) of specific sodium channel isoforms in disorders of excitability such as pain. Biological toxins have been invaluable in researching the role of ion channels in disease and some toxins that target ion channels are important therapeutics. Our preliminary data show that a toxin in the venom from the tarantula Ornithoctonus huwena inhibits TTX-sensitive neuronal sodium channels but not TTX- resistant neuronal sodium channels, cardiac sodium channels or skeletal muscle sodium channels. Furthermore, these data indicate that this toxin is more potent at blocking Nav1.6 and Nav1.7 TTX-sensitive channels than Nav1.1, 1.2 and 1.3 TTX-sensitive sodium channels. This toxin is an excellent candidate molecule on which to base the development of isoform specific sensory neuronal sodium channel blockers. This project has two related specific aims: 1. Identify the isoform specific determinants of sodium channel sensitivity to tarantula toxins. 2. Develop toxin analogs with enhanced selectivity for specific voltage-gated sodium channel isoforms. Whole-cell patch-clamp electrophysiology, mutagenesis of recombinant ion channels and peptide synthesis will be used to investigate toxin-channel interactions. The experiments proposed here will determine the feasibility of using tarantula toxins as the basis for developing novel tools to determine the roles of specific sodium channels in normal and abnormal physiology of sensory neurons. The long-term goal of this research is to develop sodium channel blockers that are useful as novel therapeutics for the treatment of neurological disorders of hyperexcitability such as chronic pain. [unreadable] [unreadable] [unreadable]